S1p receptor modulators for treating multiple sclerosis

ABSTRACT

The present invention relates uses of an S1P receptor modulator such as 2-substituted 2-amino-propane-1,3-diol or 2-amino-propanol derivatives, e.g. a compound comprising a group of formula X 
     
       
         
         
             
             
         
       
     
     for the treatment or prevention of neo-angiogenesis associated with a demyelinating disease, e.g. multiple sclerosis.

The present invention relates to the use of an S1P receptor modulator in the treatment or prevention of neo-angiogenesis associated with a demyelinating disease, e.g. multiple sclerosis.

S1 P receptor modulators are typically sphingosine analogues, such as 2-substituted 2-amino-propane-1,3-diol or 2-amino-propanol derivatives, e.g. a compound comprising a group of formula X.

Sphingosine-1 phosphate (hereinafter “S1P”) is a natural serum lipid. Presently there are eight known S1P receptors, namely S1P1 to S1P8. S1 P receptor modulators are typically sphingosine analogues, such as 2-substituted 2-amino-propane-1,3-diol or 2-amino-propanol derivatives, e.g. a compound comprising a group of formula X

wherein Z is H, C₁₋₆alkyl, C₂₋₆alkenyl, C₂₋₆alkynyl, phenyl, phenyl substituted by OH, C₁₋₆alkyl substituted by 1 to 3 substituents selected from the group consisting of halogen, C₃₋₈cycloalkyl, phenyl and phenyl substituted by OH, or CH₂—R_(4z) wherein R_(4z) is OH, acyloxy or a residue of formula (a)

wherein Z₁ is a direct bond or O, preferably O; each of R_(5z) and R_(6z), independently, is H, or C₁₋₄alkyl optionally substituted by 1, 2 or 3 halogen atoms; R_(1z) is OH, acyloxy or a residue of formula (a); and each of R_(2z) and R_(3z) independently, is H, C₁₋₄alkyl or acyl.

Group of formula X is a functional group attached as a terminal group to a moiety which may be hydrophilic or lipophilic and comprise one or more aliphatic, alicyclic, aromatic and/or heterocyclic residues, to the extent that the resulting molecule wherein at least one of Z and R_(1z) is or comprises a residue of formula (a), signals as an agonist at one of more sphingosine-1-phosphate receptor.

S1P receptor modulators are compounds which signal as agonists at one or more sphingosine-1 phosphate receptors, e.g. S1P1 to S1P8. Agonist binding to a S1P receptor may e.g. result in dissociation of intracellular heterotrimeric G-proteins into Gα-GTP and Gβγ-GTP, and/or increased phosphorylation of the agonist-occupied receptor and activation of downstream signaling pathways/kinases.

The binding affinity of S1P receptor modulators to individual human S1P receptors may be determined in following assay:

S1P receptor modulator activities of compounds are tested on the human S1P receptors S1P₁, S1P₂, S1P₃, S1P₅ and S1P₅. Functional receptor activation is assessed by quantifying compound induced GTP [γ-³⁵S] binding to membrane protein prepared from transfected CHO or RH7777 cells stably expressing the appropriate human S1P receptor. The assay technology used is SPA (scintillation proximity based assay). Briefly, DMSO dissolved compounds are serially diluted and added to SPA-bead (Amersham-Pharmacia) immobilised S1P receptor expressing membrane protein (10-20 μg/well) in the presence of 50 mM Hepes, 100 mM NaCl, 10 mM MgCl₂, 10 μM GDP, 0.1% fat free BSA and 0.2 nM GTP [γ-³⁵S] (1200 Ci/mmol). After incubation in 96 well microtiterplates at RT for 120 min, unbound GTP [γ-³⁵S] is separated by a centrifugation step. Luminescence of SPA beads triggered by membrane bound GTP [γ-³⁵S] is quantified with a TOPcount plate reader (Packard). EC₅₀s are calculated using standard curve fitting software. In this assay, the S1P receptor modulators preferably have a binding affinity to S1P receptor <50 nM.

Preferred S1P receptor modulators are e.g. compounds which in addition to their S1P binding properties also have accelerating lymphocyte homing properties, e.g. compounds which elicit a lymphopenia resulting from a re-distribution, preferably reversible, of lymphocytes from circulation to secondary lymphatic tissue, without evoking a generalized immunosuppression. Naïve cells are sequestered; CD4 and CD8 T-cells and B-cells from the blood are stimulated to migrate into lymph nodes (LN) and Peyer's patches (PP).

The lymphocyte homing property may be measured in following Blood Lymphocyte Depletion assay:

A S1P receptor modulator or the vehicle is administered orally by gavage to rats. Tail blood for hematological monitoring is obtained on day −1 to give the baseline individual values, and at 2, 6, 24, 48 and 72 hours after application. In this assay, the S1P receptor agonist or modulator depletes peripheral blood lymphocytes, e.g. by 50%, when administered at a dose of e.g. <20 mg/kg.

Examples of appropriate S1P receptor modulators are, for example:

-   -   Compounds as disclosed in EP627406A1, e.g. a compound of formula         I

-   -    wherein R₁ is a straight- or branched (C₁₂₋₂₂)chain     -   which may have in the chain a bond or a hetero atom selected         from a double bond, a triple bond, O, S, NR₆, wherein R₆ is H,         C₁₋₄alkyl, aryl-C₁₋₄alkyl, acyl or (C₁₋₄alkoxy)carbonyl, and         carbonyl, and/or         -   which may have as a substituent C₁₋₄alkoxy, C₂₋₄alkenyloxy,             C₂₋₄alkynyloxy, acyl, C₁₋₄alkylamino, C₁₋₄alkylthio,             acylamino, (C₁₋₄alkoxy)carbonyl, (C₁₋₄alkoxy)-carbonylamino,             acyloxy, (C₁₋₄alkyl)carbamoyl, nitro, halogen, amino,             hydroxyimino, hydroxy or carboxy; or

R₁ is

-   -   a phenylalkyl wherein alkyl is a straight- or branched         (C₆₋₂₀)carbon chain; or     -   a phenylalkyl wherein alkyl is a straight- or branched         (C₁₋₃₀)carbon chain wherein said phenylalkyl is substituted by     -   a straight- or branched (C₆₋₂₀)carbon chain optionally         substituted by halogen,     -   a straight- or branched (C₆₋₂₀)alkoxy chain optionally         substituted by halogen,     -   a straight- or branched (C₆₋₂₀)alkenyloxy,     -   phenyl-C₁₋₁₄alkoxy, halophenyl-C₁₋₄alkoxy,         phenyl-C₁₋₁₄alkoxy-C₁₋₁₄alkyl, phenoxy-C₁₋₄alkoxy or         phenoxy-C₁₋₄alkyl,     -   cycloalkylalkyl substituted by C₆₋₂₀alkyl,     -   heteroarylalkyl substituted by C₆₋₂₀alkyl,     -   heterocyclic C₆₋₂₀alkyl or     -   heterocyclic alkyl substituted by C₂₋₂₀alkyl,         and wherein         the alkyl moiety may have     -   in the carbon chain, a bond or a heteroatom selected from a         double bond, a triple bond, O, S, sulfinyl, sulfonyl, or NR₆,         wherein R₆ is as defined above, and     -   as a substituent C₁₋₄alkoxy, C₂₋₄alkenyloxy, C₂₋₄alkynyloxy,         aryl, C₁₋₄alkyl-amino, C₁₋₄alkylthio, acylamino,         (C₁₋₄alkoxy)carbonyl, (C₁₋₄alkoxy)carbonylamino, acyloxy,         (C₁₋₄alkyl)carbamoyl, nitro, halogen, amino, hydroxy or carboxy,         and         each of R₂, R₃, R₄ and R₅, independently, is H, C₁₋₄ alkyl or         acyl         or a pharmaceutically acceptable salt or hydrate thereof;     -   Compounds as disclosed in EP 1002792A1, e.g. a compound of         formula II

-   -    wherein m is 1 to 9 and each of R′₂, R′₃, R′₄ and R′₅,         independently, is H, C₁₋₆alkyl or acyl, or a pharmaceutically         acceptable salt or hydrate thereof;     -   Compounds as disclosed in EP0778263 A1, e.g. a compound of         formula III

-   -    wherein W is H; C₁₋₆alkyl, C₂₋₆alkenyl or C₂₋₆alkynyl;         unsubstituted or by OH substituted phenyl; R″₄O(CH₂)_(n); or         C₁₋₆alkyl substituted by 1 to 3 substituents selected from the         group consisting of halogen, C₃₋₈cycloalkyl, phenyl and phenyl         substituted by OH;         X is H or unsubstituted or substituted straight chain alkyl         having a number p of carbon atoms or unsubstituted or         substituted straight chain alkoxy having a number (p-1) of         carbon atoms, e.g. substituted by 1 to 3 substitutents selected         from the group consisting of C₁₋₆alkyl, OH, C₁₋₆alkoxy, acyloxy,         amino, C₁₋₆alkylamino, acylamino, oxo, haloC₁₋₆alkyl, halogen,         unsubstituted phenyl and phenyl substituted by 1 to 3         substituents selected from the group consisting of C₁₋₆alkyl,         OH, C₁₋₆alkoxy, acyl, acyloxy, amino, C₁₋₆alkylamino, acylamino,         haloC₁₋₆alkyl and halogen; Y is H, C₁₋₆alkyl, OH, C₁₋₆alkoxy,         acyl, acyloxy, amino, C₁₋₆alkylamino, acylamino, haloC₁₋₆alkyl         or halogen, Z₂ is a single bond or a straight chain alkylene         having a number or carbon atoms of q,         each of p and q, independently, is an integer of 1 to 20, with         the proviso of 6≦p+q≦23, m′ is 1, 2 or 3, n is 2 or 3,         each of R″₁, R^(″) ₂, R^(″) ₃ and R″₄, independently, is H,         C₁₋₄alkyl or acyl,         or a pharmaceutically acceptable salt or hydrate thereof,     -   Compounds as disclosed in WO02/18395, e.g. a compound of formula         IVa or IVb

-   -    wherein X_(a) is O, S, NR_(1s) or a group —(CH₂)_(na)—, which         group is unsubstituted or substituted by 1 to 4 halogen; n_(a)         is 1 or 2, R_(1s) is H or (C₁₋₄)alkyl, which alkyl is         unsubstituted or substituted by halogen; R_(1a) is H, OH,         (C₁₋₄)alkyl or O(C₁₋₄)alkyl wherein alkyl is unsubstituted or         substituted by 1 to 3 halogen; R_(1b) is H, OH or (C₁₋₄)alkyl,         wherein alkyl is unsubstituted or substituted by halogen; each         R_(2a) is independently selected from H or (C₁₋₄)alkyl, which         alkyl is unsubstituted or substituted by halogen; R_(3a) is H,         OH, halogen or O(C₁₋₄)alkyl wherein alkyl is unsubstituted or         substituted by halogen; and R_(3b) is H, OH, halogen,         (C₁₋₄)alkyl wherein alkyl is unsubstituted or substituted by         hydroxy, or O(C₁₋₄)alkyl wherein alkyl is unsubstituted or         substituted by halogen; Y_(a) is —CH₂—, —C(O)—, —CH(OH)—,         —C(═NOH)—, O or S, and R_(4a) is (C₄₋₁₄)alkyl or (C₄₋₁₄)alkenyl;         or a pharmaceutically acceptable salt or hydrate thereof;     -   Compounds as disclosed in WO02/06268A1, e.g. a compound of         formula V

-   -    wherein each of R_(1d) and R_(2d), independently, is H or an         amino-protecting group;         R_(3d) is hydrogen, a hydroxy-protecting group or a residue of         formula

R_(4d) is C₁₋₄alkyl; n_(d) is an integer of 1 to 6; X_(d) is ethylene, vinylene, ethynylene, a group having a formula -D-CH₂— (wherein D is carbonyl, —CH(OH)—, O, S or N), aryl or aryl substituted by up to three substitutents selected from group a as defined hereinafter; Y_(d) is single bond, C₁₋₁₀alkylene, C₁₋₁₀alkylene which is substituted by up to three substitutents selected from groups a and b, C₁₋₁₀alkylene having O or S in the middle or end of the carbon chain, or C₁₋₁₀alkylene having O or S in the middle or end of the carbon chain which is substituted by up to three substituents selected from groups a and b; R_(5d) is hydrogen, C₃₋₆cycloalkyl, aryl, heterocyclic group, C₃₋₆cycloalkyl substituted by up to three substituents selected from groups a and b, aryl substituted by up to three substituents selected from groups a and b, or heterocyclic group substituted by up to three substituents selected from groups a and b; each of R_(6d) and R_(7d), independently, is H or a substituent selected from group a; each of R_(8d) and R_(9d), independently, is H or C₁₋₄alkyl optionally substituted by halogen; <group a> is halogen, lower alkyl, halogeno lower alkyl, lower alkoxy, lower alkylthio, carboxyl, lower alkoxycarbonyl, hydroxy, lower aliphatic acyl, amino, mono-lower alkylamino, di-C₁₋₄alkylamino, acylamino, cyano or nitro; and <group b> is C₃₋₆cycloalkyl, aryl or heterocyclic group, each being optionally substituted by up to three substituents selected from group a; with the proviso that when R_(5d) is hydrogen, Y_(d) is a either a single bond or linear C₁₋₁₀ alkylene, or a pharmacologically acceptable salt, ester or hydrate thereof;

-   -   Compounds as disclosed in JP-14316985 (JP2002316985), e.g. a         compound of formula VI

-   -    wherein R_(1e), R_(2e), R_(3e), R_(4e), R_(5e), R_(6e), R_(7e),         n_(e), X_(e) and Y_(e) are as disclosed in JP-14316985;         or a pharmacologically acceptable salt, ester or hydrate         thereof;     -   Compounds as disclosed in WO03/062252A1, e.g. a compound of         formula VII

wherein Ar is phenyl or naphthyl; each of m_(g) and n₉ independently is 0 or 1; A is selected from COOH, PO₃H₂, PO₂H, SO₃H, PO(C₁₋₃alkyl)OH and 1H-tetrazol-5-yl; each of R_(1g) and R_(2g) independently is H, halogen, OH, COOH or C₁₋₄alkyl optionally substituted by halogen; R_(3g) is H or C₁₋₄alkyl optionally substituted by halogen or OH; each R_(4g) independently is halogen, or optionally halogen substituted C₁₋₄alkyl or C₁₋₃alkoxy; and each of R_(g) and M has one of the significances as indicated for B and C, respectively, in WO03/062252A1; or a pharmacologically acceptable salt, solvate or hydrate thereof;

-   -   Compounds as disclosed in WO 03/062248A2, e.g. a compound of         formula VIII

-   -    wherein Ar is phenyl or naphthyl; n is 2, 3 or 4; A is COOH,         1H-tetrazol-5-yl, PO₃H₂, PO₂H₂, —SO₃H or PO(R_(5h))OH wherein         R_(5h) is selected from C₁₋₄alkyl, hydroxyC₁₋₄alkyl, phenyl,         —CO—C₁₋₃alkoxy and —CH(OH)-phenyl wherein said phenyl or phenyl         moiety is optionally substituted;         each of R_(1h) and R_(2h) independently is H, halogen, OH, COOH,         or optionally halogeno substituted C₁₋₆alkyl or phenyl; R_(3h)         is H or C₁₋₄alkyl optionally substituted by halogen and/OH; each         R_(4h) independently is halogeno, OH, COOH, C₁₋₄alkyl,         S(O)_(0,1 or 2)C₁₋₃alkyl, C₁₋₃alkoxy, C₃₋₆cycloalkoxy, aryl or         aralkoxy, wherein the alkyl portions may optionally be         substituted by 1-3 halogens; and each of R_(h) and M has one of         the significances as indicated for B and C, respectively, in         WO03/062248A2         or a pharmacologically acceptable salt, solvate or hydrate         thereof.     -   Compounds as disclosed in WO 04/103306A, WO 05/000833, WO         05/103309 or WO 05/113330, e.g. compounds of formula IXa or IXb

wherein A_(k) is COOR_(5k), OPO(OR_(5k))₂, PO(OR_(5k))₂, SO₂OR_(5k), POR_(5k)OR_(5k) or 1H-tetrazol-5-yl, R_(5k) being H or C₁₋₆alkyl; W_(k) is a bond, C₁₋₃alkylene or C₂₋₃alkenylene; Y_(k) is C₆₋₁₀aryl or C₃₋₉heteroaryl, optionally substituted by 1 to 3 radicals selected from halogene, OH, NO₂, C₁₋₆alkyl, C₁₋₆alkoxy; halo-substituted C₁₋₆alkyl and halo-substituted C₁₋₆alkoxy; Z_(k) is a heterocyclic group as indicated in WO 04/103306A, e.g. azetidine; R_(1k) is C₆₋₁₀aryl or C₃₋₉heteroaryl, optionally substituted by C₁₋₆alkyl, C₆₋₁₀aryl, C₆₋₁₀arylC₁₋₄alkyl, C₃₋₉heteroaryl, C₃₋₉heteroarylC₁₋₄alkyl, C₃₋₈cycloalkyl, C₃₋₈cycloalkylC₁₋₄alkyl, C₃₋₈heterocycloalkyl or C₃₋₈heterocycloalkylC₁₋₄alkyl; wherein any aryl, heteroaryl, cycloalkyl or heterocycloalkyl of R_(1k) may be substituted by 1 to 5 groups selected from halogen, C₁₋₆alkyl, C₁₋₆alkoxy and halo substituted-C₁₋₆alkyl or —C₁₋₆alkoxy; R_(2k) is H, C₁₋₆alkyl, halo substituted C₁₋₆alkyl, C₂₋₆alkenyl or C₂₋₆alkynyl: and each of R_(3k) or R_(4k), independently, is H, halogen, OH, C₁₋₆alkyl, C₁₋₆alkoxy or halo substituted C₁₋₆alkyl or C₁₋₆alkoxy; and the N-oxide derivatives thereof or prodrugs thereof, or a pharmacologically acceptable salt, solvate or hydrate thereof.

The compounds of formulae I to IXb may exist in free or salt form. Examples of pharmaceutically acceptable salts of the compounds of the formulae I to VI include salts with inorganic acids, such as hydrochloride, hydrobromide and sulfate, salts with organic acids, such as acetate, fumarate, maleate, benzoate, citrate, malate, methanesulfonate and benzenesulfonate salts, or, when appropriate, salts with metals such as sodium, potassium, calcium and aluminium, salts with amines, such as triethylamine and salts with dibasic amino acids, such as lysine. The compounds and salts of the combination of the present invention encompass hydrate and solvate forms.

Acyl as indicated above may be a residue R_(y)—CO— wherein R_(y) is C₁₋₆alkyl, C₃₋₆cycloalkyl, phenyl or phenyl-C₁₋₄alkyl. Unless otherwise stated, alkyl, alkoxy, alkenyl or alkynyl may be straight or branched.

Aryl may be phenyl or naphthyl, preferably phenyl.

When in the compounds of formula I the carbon chain as R₁ is substituted, it is preferably substituted by halogen, nitro, amino, hydroxy or carboxy. When the carbon chain is interrupted by an optionally substituted phenylene, the carbon chain is preferably unsubstituted. When the phenylene moiety is substituted, it is preferably substituted by halogen, nitro, amino, methoxy, hydroxy or carboxy.

Preferred compounds of formula I are those wherein R₁ is C₁₃₋₂₀alkyl, optionally substituted by nitro, halogen, amino, hydroxy or carboxy, and, more preferably those wherein R₁ is phenylalkyl substituted by C₆₋₁₄-alkyl chain optionally substituted by halogen and the alkyl moiety is a C₁₋₆alkyl optionally substituted by hydroxy. More preferably, R₁ is phenyl-C₁₋₆alkyl substituted on the phenyl by a straight or branched, preferably straight, C₆₋₁₄alkyl chain. The C₆₋₁₄alkyl chain may be in ortho, meta or para, preferably in para.

Preferably each of R₂ to R₅ is H.

In the above formula of V “heterocyclic group” represents a 5- to 7 membered heterocyclic group having 1 to 3 heteroatoms selected from S, O and N. Examples of such heterocyclic groups include the heteroaryl groups indicated above, and heterocyclic compounds corresponding to partially or completely hydrogenated heteroaryl groups, e.g. furyl, thienyl, pyrrolyl, azepinyl, pyrazolyl, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, 1,2,3-oxadiazolyl, triazolyl, tetrazolyl, thiadiazolyl, pyranyl, pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, tetrahydropyranyl, morpholinyl, thiomorpholinyl, pyrrolidinyl, pyrrolyl, imidazolidinyl, pyrazolidinyl, piperidinyl, piperazinyl, oxazolidinyl, isoxazolidinyl, thiazolidinyl or pyrazolidinyl. Preferred heterocyclic groups are 5- or 6-membered heteroaryl groups and the most preferred heteocyclic group is a morpholinyl, thiomorpholinyl or piperidinyl group.

A preferred compound of formula I is 2-amino-2-tetradecyl-1,3-propanediol. A particularly preferred S1P receptor agonist of formula I is FTY720, i.e. 2-amino-2-[2-(4-octylphenyl)ethyl]propane-1,3-diol in free form or in a pharmaceutically acceptable salt form (referred to hereinafter as Compound A), e.g. the hydrochloride salt, as shown:

A preferred compound of formula II is the one wherein each of R′₂ to R^(′) ₅ is H and m is 4, i.e. 2-amino-2-{2-[4-(1-oxo-5-phenylpentyl)phenyl]ethyl}propane-1,3-diol, in free form or in pharmaceutically acceptable salt form (referred to hereinafter as Compound B), e.g the hydrochloride.

A preferred compound of formula III is the one wherein W is CH₃, each of R″₁ to R″₃ is H, Z₂ is ethylene, X is heptyloxy and Y is H, i.e. 2-amino-4-(4-heptyloxyphenyl)-2-methyl-butanol, in free form or in pharmaceutically acceptable salt form (referred to hereinafter as Compound C), e.g. the hydrochloride. The R-enantiomer is particularly preferred.

Compounds may be in phosphorylated form. A preferred compound of formula IVa is the FTY720-phosphate (R_(2a) is H, R_(3a) is OH, X_(a) is O, R_(1a) and R_(1b) are OH). A preferred compound of formula IVb is the Compound C-phosphate (R_(2a) is H, R_(3b) is OH, X_(a) is O, R_(1a) and R_(1b) are OH, Y_(a) is O and R_(4a) is heptyl). A preferred compound of formula V is Compound B-phosphate.

A preferred compound of formula VI is (2R)-2-amino-4-[3-(4-cyclohexyloxybutyl)-benzo[b]thien-6-yl]-2-methylbutan-1-ol.

A preferred compound of formula IXa is e.g. 1-{4-[1-(4-cyclohexyl-3-trifluoromethyl-benzyloxyimino)-ethyl]-2-ethyl-benzyl}-azetidine-3-carboxylic acid, or a prodrug thereof.

S1P receptor agonists or modulators are known as having immunosuppressive properties or anti-angiogenic properties in the treatment of tumors, e.g. as disclosed in EP627406A1, WO 04/103306, WO 05/000833, WO 05/103309, WO 05/113330 or WO 03/097028.

Multiple sclerosis (MS) is an immune-mediated disease of the central nervous system with chronic inflammatory demyelination leading to progressive decline of motor and sensory functions and permanent disability. The therapy of multiple sclerosis is only partially effective, and in most cases only offers a short delay in disease progression despite anti-inflammatory and immunosuppressive treatment. Accordingly, there is a need for agents which are effective in the inhibition or treatment of demyelinating diseases, e.g. multiple sclerosis or Guillain-Barré syndrome, including reduction of, alleviation of, stabilization of or relief from the symptoms which affect the organism.

Characteristic pathological features of demyelinating diseases include inflammation, demyelination and axonal and oligodendrocyte loss. In addition lesions can also have a significant vascular component. A firm link has recently been established between chronic inflammation and angiogenesis and neovascularization seems to have a significant role in the progression of disease.

It has now been found that S1P receptor modulators have an inhibitory effect on neo-angiogenesis associated with demyelinating diseases, e.g. MS.

In a series of further specific or alternative embodiments, the present invention provides:

-   1.1 A method for preventing, inhibiting or treating neo-angiogenesis     associated with a demyelinating disease, e.g. MS, in a subject in     need thereof, comprising administering to said subject a     therapeutically effective amount of an S1P receptor modulator, e.g.     a compound of formulae I to IXb. -   1.2 A method for alleviating or delaying progression of the symptoms     of a demyelinating disease, e.g. multiple sclerosis or     Guillain-Barré syndrome, in a subject in need thereof, in which     method neo-angiogenesis associated with said disease is prevented or     inhibited, comprising administering to said subject a     therapeutically effective amount of an S1P receptor modulator, e.g.     a compound of formulae I to IXb. -   1.3 A method for reducing or preventing or alleviating relapses in a     demyelinating disease, e.g. multiple sclerosis or Guillain-Barré     syndrome, in a subject in need thereof, in which method     neo-angiogenesis associated with said disease is prevented or     inhibited, comprising administering to said subject a     therapeutically effective amount of an S1P receptor modulator, e.g.     a compound of formulae I to IXb. -   1.4 A method for slowing progression of a demyelinating disease,     e.g. multiple sclerosis or Guillain-Barré syndrome, in a subject     being in a relapsing-remitting phase of the disease, in which method     neo-angiogenesis associated with said disease is prevented or     inhibited, comprising administering to said subject a     therapeutically effective amount of an S1P receptor modulator, e.g.     a compound of formulae I to XIb. -   1.5 A method as indicated above, wherein the S1P receptor modulator     is administered intermittently.     -   For example, the S1P receptor modulator may be administered to         the subject every 2^(nd) or 3^(rd) day or once a week. -   2. A pharmaceutical composition for use in any one of the methods     1.1 to 1.5, comprising an S1P receptor modulator, e.g. a compound of     formulae I to IXb as defined hereinabove, together with one or more     pharmaceutically acceptable diluents or carriers therefor. -   3. An S1P receptor modulator, e.g a compound of formula I to IXb as     defined herein above, for use in any one of the methods 1.1 to 1.5. -   4 An S1P receptor modulator, e.g. a compound of formulae I to IXb as     defined herein above, for use in the preparation of a medicament for     use in any one of the methods 1.1 to 1.5.

Clinicians usually categorize patients having MS into four types of disease patterns:

-   -   Relapsing-remitting (RR-MS): Discrete motor, sensory, cerebellar         or visual attacks that occur over 1-2 weeks and often resolve         over 1-2 months. Some patients accrue disability with each         episode, yet remain clinically stable between relapses. About         85% of patients initially experience the RR form of MS, but         within 10 years about half will develop the secondary         progressive form.     -   Secondary-progressive (SP-MS): Initially RR followed by         gradually increasing disability, with or without relapses. Major         irreversible disabilities appear most often during SP.     -   Primary-progressive (PP-MS): Progression disease course from         onset without any relapses or remissions, affecting about 15% of         MS patients.     -   Progressive-relapsing (PR-MS): Progressive disease from onset         with clear acute relapses; periods between relapses         characterized by continuing progression.

Accordingly, the S1P receptor modulators, e.g. a compound of formulae I to IXb as defined hereinabove, may be useful in the treatment of one or more of Relapsing-remitting (RR-MS), Secondary-progressive (SP-MS), Primary-progressive (PP-MS) and Progressive-relapsing (PR-MS).

In particular, the S1P receptor modulators as described herein, e.g. FTY720, i.e. 2-amino-2-[2-(4-octylphenyl)ethyl]propane-1,3-dio, are useful for treating PP-MS.

Utility of the S1P receptor modulators, e.g. the S1P receptor modulators comprising a group of formula X, in preventing or treating neo-angiogenesis associated with a demyelinating disease as hereinabove specified, may be demonstrated in animal test methods as well as in clinic, for example in accordance with the methods hereinafter described.

In Vivo: Relapsing Experimental Autoimmune Encephalomyelitis (EAE)

Disease is induced in female Lewis rats by immunization with guinea pig spinal cord tissue emulsified in complete Freund's adjuvant. This results in an acute disease within 11 days, followed by an almost complete remission around day 16 and a relapse at around days 26. On day 26 rats are thoracectomized after having been deeply anesthetized with Isoflurane (3%, 20 L/min) and perfused through the left ventricle of the heart. The left ventricle is punctured with a 19 gauge needle from a winged infusion set (SV-19BLK; Termudo, Elkton, Md.), which is connected to an airtight pressurized syringe containing the rinsing solution (NaCl 0.9% with 250,000 U/I heparin at 35° C.). The right atrium is punctured to provide outflow, and the perfusate is infused under a precise controlled pressure of 120 mm Hg. The perfusion is continued for 5 min (at a constant rate of 20 ml/min) followed by a pre-fixation solution (2% performaldehyde in PBS at 35° C.). Finally, up to 30 ml of polyurethane resin (PUII4; Vasqtec, Zúrich, Switzerland) is infused at the same rate. After 48 h, the resin-filled brain and spinal cord are excised from the animal and the soft tissue removed by maceration in 7.5% KOH during 24 hr at 50° C. The casts are then thoroughly cleaned with and stored in distilled water before drying by lyophilization. These vascular casts are quantitated using micro computer tomography.

In this assay, a S1P1 receptor modulator, e.g. Compound A significantly blocks disease-associated neo-angiogenesis when administered to the animals at a dose of from 0.1 to 20 mg/kg p.o. For example, Compound A, in the hydrochloride salt form, fully blocks disease-associated angiogenesis and completely inhibits the relapse phases when administered daily at a dose of 0.3 mg/kg p.o. The same effect is obtained when Compound A, in the hydrochloride salt form, is administered p.o. at 0.3 mg/kg every 2^(nd) or 3^(rd) day or once a week.

C. Clinical Trial

Investigation of clinical benefit of a S1P receptor agonist, e.g. a compound of formula I, e.g. Compound A.

20 patients with relapsing-remitting MS receive said compound at a daily dosage of 0.5, 1.25 or 2.5 mg p.o. The general clinical state of the patient is investigated weekly by physical and laboratory examination. Disease state and changes in disease progression are assessed every 2 months by radiological examination (MRI) and physical examination. Initially patients receive treatment for 2 to 6 months. Thereafter, they remain on treatment for as long as their disease does not progress and the drug is satisfactorily tolerated.

Main variables for evaluation: Safety (adverse events), standard serum biochemistry and hematology, magnetic resonance imaging (MRI).

Daily dosages required in practicing the method of the present invention when a S1P receptor modulator alone is used will vary depending upon, for example, the compound used, the host, the mode of administration and the severity of the condition to be treated. A preferred daily dosage range is about from 0.1 to 100 mg as a single dose or in divided doses. Suitable daily dosages for patients are on the order of from e.g. 0.1 to 50 mg p.o. The S1P receptor modulator may be administered by any conventional route, in particular enterally, e.g. orally, e.g. in the form of tablets, capsules, drink solutions, nasally, pulmonary (by inhalation) or parenterally, e.g. in the form of injectable solutions or suspensions. Suitable unit dosage forms for oral administration comprise from ca. 0.1 to 30 mg, usually 0.25 to 30 mg S1P receptor modulator, together with one or more pharmaceutically acceptable diluents or carriers therefore. As already mentioned, the S1Preceptor modulator, e.g. Compound A, may alternatively be administered intermittently, e.g. at a dose of 0.5 to 30 mg every other day or once a week.

According to another embodiment of the invention, the S1P receptor modulator may be administered as the sole active ingredient or in conjunction with, e.g. as an adjuvant to, a VEGF-receptor antagonist.

Examples of suitable VEGF-receptor antagonist include e.g. compounds, proteins or antibodies which inhibit the VEGF receptor tyrosine kinase, inhibit a VEGF receptor or bind to VEGF, and are e.g. in particular those compounds, proteins or monoclonal antibodies generically and specifically disclosed in WO 98/35958, e.g. 1-(4-chloroanilino)-4-(4-pyridylmethyl)phthalazine or a pharmaceutically acceptable salt thereof, e.g. the succinate, in WO 00/27820, e.g. a N-aryl(thio) anthranilic acid amide derivative e.g. 2-[(4-pyridyl)methyl]amino-N-[3-methoxy-5-(trifluoromethyl)phenyl]benzamide or 2-[(1-oxido-4-pyridyl)methyl]amino-N-[3-trifluoromethylphenyl]benzamide, or in WO 00/09495, WO 00/59509, WO 98/11223, WO 00/27819, WO 01/55114, WO 01/58899 and EP 0 769 947; those as described by M. Prewett et al in Cancer Research 59 (1999) 5209-5218, by F. Yuan et al in Proc. Natl. Acad. Sci. USA, vol. 93, pp. 14765-14770, December 1996, by Z. Zhu et al in Cancer Res. 58, 1998, 3209-3214, and by J. Mordenti et al in Toxicologic Pathology, Vol. 27, no. 1, pp 14-21, 1999; in WO 00/37502 and WO 94/10202; Angiostatin™, described by M. S. O'Reilly et al, Cell 79, 1994, 315-328; Endostatin™, described by M. S. O'Reilly et al, Cell 88, 1997, 277-285; anthranilic acid amides; ZD4190; ZD6474; SU5416; SU6668; or anti-VEGF antibodies or anti-VEGF receptor antibodies, e.g. RhuMab.

4-Pyridylmethyl-phthalazine derivatives are e.g. preferred inhibitors of VEGF receptor tyrosine kinase. Such derivatives and their preparation, pharmaceutical formulations thereof and methods of making such compounds are described in WO00/59509, EP02/04892, WO01/10859 and, in particular, in U.S. Pat. No. 6,258,812, which are here incorporated by reference.

Where the S1P receptor modulator is administered in conjunction with a VEGF-receptor antagonist, dosages of the co-administered VEGF-receptor agonist will of course vary depending on the type of co-drug employed, e.g. whether it is a steroid or a calcineurin inhibitor, on the specific drug employed, on the condition being treated and so forth. In accordance with the foregoing the present invention provides in a yet further aspect:

-   5. A method as defined above comprising co-administration, e.g.     concomitantly or in sequence, of a therapeutically effective     non-toxic amount of a S1P receptor modulator and a VEGF-receptor     antagonist, e.g. as indicated above. -   6. A pharmaceutical combination, e.g. a kit, comprising a) a first     agent which is a S1P receptor modulator as disclosed herein, in free     form or in pharmaceutically acceptable salt form, and b) a     VEGF-receptor antagonist, e.g. as indicated above. The kit may     comprise instructions for its administration.

The terms “co-administration” or “combined administration” or the like as utilized herein are meant to encompass administration of the selected therapeutic agents to a single patient, and are intended to include treatment regimens in which the agents are not necessarily administered by the same route of administration or at the same time.

The term “pharmaceutical combination” as used herein means a product that results from the mixing or combining of more than one active ingredient and includes both fixed and non-fixed combinations of the active ingredients. The term “fixed combination” means that the active ingredients, e.g. a S1P receptor modulator and a VEGF-receptor antagonist, are both administered to a patient simultaneously in the form of a single entity or dosage. The term “non-fixed combination” means that the active ingredients, e.g. a S1P receptor modulator and a VEGF-receptor antagonist, are both administered to a patient as separate entities either simultaneously, concurrently or sequentially with no specific time limits, wherein such administration provides therapeutically effective levels of the 2 compounds in the body of the patient. 

1-3. (canceled)
 4. A method according to claim 7, wherein the medicament is co-administered, e.g. concomitantly or in sequence, with a VEGF-receptor antagonist.
 5. A pharmaceutical composition for use in the method according to claim 7, comprising an S1P receptor modulator together with one or more pharmaceutically acceptable diluents or carriers therefor.
 6. A pharmaceutical combination, e.g. a kit, comprising a) a first agent which is a S1P receptor modulator, in free form or in pharmaceutically acceptable salt form, and b) a VEGF-receptor antagonist.
 7. A method for preventing, inhibiting or treating neo-angiogenesis associated with a demyelinating disease in a subject in need thereof, comprising administering to said subject a therapeutically effective amount of an S1P receptor modulator.
 8. A method according to claim 7, wherein the demyelinating disease is multiple sclerosis.
 9. A method according to claim 8, wherein the S1P receptor modulator is administered intermittently.
 10. A method according to claim 14, wherein the S1P receptor modulator or agonist is 2-amino-2-[2-(4-octylphenyl)ethyl]propane-1,3-diol, 2-amino-2-[4-(3-benzyloxyphenoxy)-2-chlorophenyl]ethyl-1,3-propane-diol, or 1-{4-[1-(4-cyclohexyl-3-trifluoromethyl-benzyloxyimino)-ethyl]-2-ethyl-benzyl}-azetidine-3-carboxylic acid, in free form or in a pharmaceutically acceptable salt form.
 11. A method according to claim 10, wherein the S1P receptor modulator is 2-amino-2-[2-(4-octylphenyl)ethyl]propane-1,3-diol in free form or in a pharmaceutically acceptable salt form.
 12. A method according to claim 7, wherein the demyelinating disease is primary progressive multiple sclerosis (PP-MS).
 13. A method according to claim 7, wherein the S1P receptor modulator comprises a group of formulae I to IXb.
 14. A method according to claim 7, wherein the S1P receptor modulator comprises a group of formula X:

wherein Z is H, C₁₋₆alkyl, C₂₋₆alkenyl, C₂₋₆alkynyl, phenyl, phenyl substituted by OH, C₁₋₆alkyl substituted by 1 to 3 substituents selected from the group consisting of halogen, C₃₋₈cycloalkyl, phenyl and phenyl substituted by OH, or CH₂—R_(4z) wherein R_(4z) is OH, acyloxy or a residue of formula (a)

wherein Z₁ is a direct bond or O, preferably O; each of R_(5z) and R_(6z), independently, is H, or C₁₋₄alkyl optionally substituted by 1, 2 or 3 halogen atoms; R_(1z) is OH, acyloxy or a residue of formula (a); and each of R_(2z) and R_(3z) independently, is H, C₁₋₄alkyl or acyl. 